The aim of the proposed research is to study the nature, disposition and intracellular origin of cell surface sialoglycoproteins. First, the sialic acid - containing components present in isolated subcellular fractions of cytosol, endoplasmic reticulum (rough microsomes), Golgi and plasma membrane of rat liver will be identified using a radiochemical labeling technique specific for sialic acid residues. Fractions such as the rough microsomes and Golgi will be subfractionated into content (i.e., soluble) proteins and purified membranes. The sialoglycoproteins in each compartment will be further characterized as to number, size and relative sialic acid content by sodium dodecyl sulfate polyacrylamide gel electrophoretic analysis. In addition, the presence of other sugars and chemical (e.g., N-terminal, peptide map, amino acid analysis) and/or immunological similarity among sialoglycoproteins in different cellular compartments will be examined. Next, the orientation and disposition (i.e., inside, outside or transmembrane) of these components in the various membrane vesicle preparations will be studied using a combination of vectorial labeling (enzymatic iodination), exposure to hydrolases (neuraminidase, trypsin, pronase) and sialic acid labeling. Finally, kinetic experiments will be conducted in order to trace the intracellular biogenetic pathway of the cell surface sialoglycoproteins. Labeled amino acids and specific monosaccharides will be injected into intact animals and perfused liver in a "pulse-chase" fashion followed by subcellular fractionation at various times and analysis of the rates at which the compartments (and individual components of them) are labeled. Again sialic acid labeling will be used to identify the sialoglycoproteins.